1. Field of the Invention
This invention relates to isolated DNA fragments encoding insulin-like growth factors I and II (IGF-I, IGF-II) isolated from the rainbow trout, Oncorhynchus mykiss, and to nucleotide probes and amplification primers specific for rainbow trout IGF-I and IGF-II, expression vectors containing the DNA encoding rainbow trout IGF-I and IGF-II, and to recombinantly produced rainbow trout IGF-I and IGF-II proteins. The invention also relates to antibodies directed to the recombinantly produced IGF-I and IGF-II proteins, and to kits for the detection of these proteins. The invention further relates to methods of treating various fish species using either recombinantly produced rainbow trout IGF-I and IGF-II proteins, or using DNA constructs containing the rainbow trout IGF-I- and IGF-II-encoding fragments.
2. Background of the Invention
The insulin gene family is an ancient and highly diverse group which includes insulin and insulin-like growth factor I (IGF-I) from a variety of species, mammalian insulin-like growth factor II (IGF-II), relaxin, insect prothoracicotrophic hormone (PTTH), and molluscan insulin-related peptide (MIP) (Blundell et al, (1980) Nature (London) 287, 781-787; Smit et al, (1989) Nature (London) 331, 535-538). IGFs are mitogenic peptide hormones that play an important role in the growth and differentiation of vertebrates (Froesh, E. R. (1983) in Insulin-like growth factors/somatomedins, ed. Spencer, M. (de Gruyter, New York), pp. 13-29).
Both insulin and IGFs are translated as prepropeptides, which can be divided into an NH.sub.2 -terminal prepeptide, a B-domain, a C-domain, and an A-domain. IGFs consist of an additional D-domain and COOH-terminal E-domain trailer peptide. The IGF prepeptide leader and trailer peptide are proteolytically removed, as is the insulin prepeptide leader. However, unlike IGF's, insulin is further processed by the removal of the internal C polypeptide domain.
IGF-I and IGF-II share approximately 50% homology. The mature form of mammalian IGF-I is a basic protein consisting of 70 amino acid residues. Two forms of IGF-I have been identified, which share approximately 95% homology. However, these two forms appear to have different biological roles. Liver, under the influence of pituitary growth hormone, is the primary site of IGF-I production. The mature form of mammalian IGF-II is a neutral protein consisting of 67 amino acid residues and is produced primarily in the liver, under the control of placental lactogen (Gray et al, (1987) DNA 6, 283-295) during prenatal development.
IGF-I and IGF-II are ubiquitous and essential regulators of cell division and vertebrate growth, and are thought to play a role in various stages of development. IGF-I is thought to be more significant in adult growth and development, while IGF-II has generally been considered a fetal form of IGF-I. Elevated levels of both IGF-I and IGF-II have been correlated to substantial increases in growth, including both mass and length, in a number of animal species.
IGF-I has been identified, either at the protein or nucleic acid level in a number of non-mammalian vertebrates including chicken (Dawe et al, (1988) J. Endocrinol. 177, 173-181), Xenopus laevis (Kajimoto et al, (1990) Mol. Endocrinol. 4, 217-225), coho salmon (Cao et al, (1989) Mol. Endocrinology 3, 2005-2010), and atlantic hagfish (Nagamatsu et al, (1991) J. Biol. Chem. 226, 2397-2404). IGF-I isolated from coho salmon and mammalian IGF-I share only an 80% homology at the amino acid level. IGF-II has not been identified in a fish species prior to the present invention, nor have the mRNA levels of IGF-I or II been determined in any fish species.
To date, there has been no efficient method for treating fish with recombinantly produced IGFs, or for effecting elevated levels of IGFs in fish species using DNA constructs containing IGF genes isolated from fish. Such treatment would allow for enhanced growth of fish, which would be highly useful in the aquaculture industry.
Therefore, in view of the lack of effective methods for treating fish with IGFs to enhance growth characteristics, it should be apparent that there still exists a need in the art for such methods.